The present invention relates to luminescent methods and compositions for detecting blood using the fluorescin to fluorescein reaction.
There are various method for detecting latent, blood-based fingerprints, palm prints and footprints at, for example, a crime scene. One such technique relies on chemiluminescence of LUMINOL (5-amino-2,3-dihydro-1,4-phthalazine) which is sprayed on the area suspected of containing blood. Oxidation of the LUMINOL in the presence of iron and peroxides found in blood is accompanied by emission of light that lasts for several seconds. A drawback of using LUMINOL is that the detection must be done in the dark to detect the emission of light, the chemiluminescence lasts for only several seconds and is not suitable such as to obtain results from a black substrate.
More recently, criminalists and crime scene investigators have used the reaction of fluorescin to fluorescein to detect the presence and pattern of bloodstains. Such a technique is as described in Cheeseman, U.S. Pat. No. 5,976,886 issued Nov. 2, 1999 the disclosure of which is hereby incorporated by reference. In this patent there is described a method for detecting latent bloodstains by applying a fluorescin solution thickened by addition of a gum compound followed by a dilute hydrogen peroxide solution to enhance the conversion of fluorescin to fluorescein and illuminating the area with UV light causing the fluorescein to fluoresce to expose the print for capture such as by photography. Te Cheeseman recipe uses water as the solvent. This stems from the original waterbased fluorescin work by Macieri and Monk. A drawback of this technique is that the fluorescin reagent according to this reference is caustic, water-soluble and has a short shelf life. The caustic nature of the solution can permanently damage the substrate being tested such as furniture, aluminum window frames and the like and may be hazardous to skin contact. The water solubility tends to result in loss of ridge detail for, for example, a fingerprint on a vertical surface such as a wall. Preserving ridge detail is crucial in matching a latent fingerprint, palm print or the like. The short shelf live requires the solution to be made relatively fresh and would undoubtedly lead to waste as aged solutions wold have to be discarded. Further there is risk that an aged solution may be used perhaps jeopardizing the collection of evidence.
Synthesis of fluorescin solutions in formic acid has been attempted, however formic acid is toxic, the reagents described are more expensive and ridge detail would probably be lost in the formic acid formulation.
There is a need for a method, solutions and kit for detection of blood stains which is non-toxic, will not damage the surface being examined, which has a longer shelf life and which provides for the fluorescin to fluorescein reaction in a less caustic solution and which is believed to result in greater detail being exhibited as well as providing for longer exhibition times for the fluorescing results. The kit preferably includes components which can be pre-formulated and wherein such formulation provides for preservation of ridge detail of the print.
Toward this end a solution and method are set forth for detecting blood stains, which preserves ridge detail, which is less caustic and less toxic, which provides for longer reaction exhibition and which substantially overcomes the drawbacks noted above.
Accordingly, the method includes applying to the area suspected of containing blood a reactant solution containing either fluorescein or dichlorofluorescein solubilized in acetic acid and ethanol with a metal reducing agent such as, for example, zinc, magnesium or aluminum. The reduction to fluorescin can be completed by one of two methods: cold solubilization or reflux solubilization. In room temperature solubilization, fluorescein or dichlorofluorescein is added to acetic acid and ethanol in a vial with a powdered metal such as zinc. The solution is lightly shaken or warmed until the yellowish color turns-clear. In the reflux solubilization, the solution should be refluxed over mossy zinc for about two hours to reduce the fluorescein, which has a yellowish color, to fluorescin which is a substantially clear solution. A buffering agent can be added to buffer the pH of the refluxed solution to a pH of between 8 and 10 and preferably to a pH of approximately 9 to increase brightness. The buffering agent is preferably an organic a mine buffer such as 2-amino-2-methyl-1-propanol in ethanol is added to the reactant solution until said solution has a neutral or slightly basic pH, e.g. a pH of approximately 9. When shelf life of the mixed solution is critical, the buffering step is ignored to increase solution stability.
Where print detail is critical on shiny surfaces that do not wet, there are two methods to decrease surface tension and the resulting spotted deposition. The surface can be pre-sprayed or dipped with/in a thin polymer solution and dried. The surface tension is lowered and the solutions will wet better producing better ridge detail. In the second method to reduce spots, a small amount of non-ionic surfactant is sprayed on the surface. These surfactants include fluoro-surfactants that can be sprayed at very low concentrations.
In one embodiment the reactant includes a Compound A of dry fluorescein with granular zinc or other metal reducing agent such as calcium, aluminum or magnesium. Since the presence of iron will poison the reduction, the reducing agent should be free of iron. Prior to application on the area suspected to have a blood stain, acetic acid and ethanol are added to solubilize compound A and mixed such that the reducing agent reduces the fluorescein to fluorescin. Reduction causes the solution to turn from a yellowish color to clear. The clear, fluorescin reactant solution is nebulized onto the suspected area.
The fluorescin of the reactant oxidizes with the blood to produce in those areas where blood is present, fluorescein.
To enhance the reaction, a Compound B oxidizing agent such as sodium metaborate, monosoduim metaborate, sodium borate or the like in a dry form or liquid sodium metaborate, monosoduim metaborate or sodium borate in alcohol or water or hydrogen peroxide. For example Compound B may include hydrogen peroxide which can be a 2-3% solution in water or a 3-5% in acetone from a 30% stock solution. Each of these oxidizing agents is used to produce an oxidizing solution, or produce hydrogen peroxide, or directly oxidize the blood on contact. Sodium metaborate has the advantage over a hydrogen peroxide solution in that sodium metaborate has a longer shelf life, is suitable for use in a kit form and provides a more controlled supply of oxidizer, or rate of peroxide formation when compared to spraying hydrogen peroxide directly.
Afer application of the fluorescin, the oxidizer is dusted or nebulized over he suspected area.
In response to UV light or blue light, e.g. wavelengths of approximately between 400 and 500 nm, the reacted fluorescein fluoresces for photographic or digital image collection of the blood print.
The compositions according to the present invention includes the synthesized and refluxed fluorescin/dichlorofluorescin reactant solution in acetic acid and ethanol which can be buffered to substantially neutral or only just basic as well as the use of the polymer and oxidant described above. The reactant solution can be provided in a kit, whereas, where the oxidizer is a hydrogen peroxide solution, should be made fresh. Where the oxidizer is a dry powder having a greater shelf life, it can be provided in a sealed container for use in the kit form. The kit would then be composed of a powdered fluorescent dye indicator, a powdered or solution of oxidizer and a bottle of ethanol.
Thus it is an object of the present invention to provide a method for detection of blood on a substrate using the fluorescin to fluorescein reaction wherein the reactant solution is not caustic, easy to mix, and preserving of ridge detail.
It is a further object to provide a reactant solution wherein the reactant is synthesized in acid and then may be buffered to the desired pH.
It is a further object to provide a reactant solution that has a greater shelf life that compositions heretofore utilized.
It is a further object to produce a composition that results in an intense and relatively persistent luminescence.
It is another of the present invention to provide a method for detection of blood an a substrate using the fluorescin to fluorescein reaction wherein the reactant solution is not caustic.
It is still a further object to provide compositions and method which are well suited to being provided in a kit form.